The long-term objectives of this proposal are to design and develop novel strategies for regulating the development and progression of atherosclerosis and to provide new insights into the application of target specific hammerhead ribozymes for inhibiting gene expression. Our ultimate goal will be to develop human gene therapy strategies for alleviating atherosclerosis, which is responsible for most clinical manifestations of coronary artery disease and ischemic stroke. Atherosclerosis is recognized as an inflammatory disease, with the presence of LDL in the intima (subendothelial matrix) leading to inflammatory responses, oxidative stress, and plaque formation. Our laboratory has developed a genetically defined mouse model of atherosclerosis (LDb mice), which has a plasma lipoprotein profile that resembles humans with hyperlipidemia and results in diet-independent atherosclerosis. The etiology of atherosclerosis in this model mimics that of humans. In this application, we plan to develop novel strategies using minimal and tertiary stabilized hammerhead ribozymes to target two genes (apolipoprotein B mRNA and lipoprotein-associated phospholipase A2) simultaneously and will test the effect of this strategy on atherosclerosis development in LDb mice. We hypothesize that inhibiting gene expression of apoB and Lp-PLA2 mRNA will decrease oxidation of LDL, reduce the inflammatory response, and inhibit atherosclerosis development in LDb mice. We will use the double-stranded chimeric adeno-associated viral vector AAV2/8 to deliver these hammerhead ribozyme molecules to mice and will examine the persistence of both vectors and ribozyme gene expression in vivo. This study will allow us to understand the molecular mechanisms used by ribozymes to regulate gene expression and the ability of these ribozymes to inhibit atherosclerosis development. In summary, this study will provide a novel therapeutic treatment for coronary atherosclerosis and provide evidence of the efficiency and efficacy of RNA based molecules as potential therapeutic agents.